Zika virus (ZIKV) has recently been declared a new global threat and an emerging pathogen in the USA. Common features of ZIKV infection include little to no symptoms for the majority of infected adults, but debilitating neurological symptoms such as Guillain-Barr syndrome for a few infected patients. While cause-and-effect has not been proven, there is a startling and troubling correlation between ZIKV infection in pregnant mothers and microcephaly in their newborn infants. There is currently no pharmacological protection for humans against mosquito-borne flaviviral diseases. The major goal of this proposal is to develop the mechanistic understanding of the ZIKV pathology in the fetus brain. This understanding can ultimately be developed into drugs to treat ZIKV disease. It becomes now obvious that there is complex, well-balanced mutualism between the human cells and the virus, which reciprocally affect each other through multiple, albeit insufficiently understood, molecular, cellular and biochemical mechanisms. It is exceedingly difficult, if ever possible, to understand these mechanisms by using the irrelevant cell types. In contrast with many others, our study is focused on neural precursors, the cell type infected by ZIKV in the fetus brain. A major strength of our application lies in the highly relevant expertise of our team of biochemists, bioinformaticians, neurobiologists, and virologists from the laboratories of Drs. Terskikh, Cieplak and Strongin (SBP Discovery), together with Dr. Shresta (LJI). There are two overarching hypotheses that direct our research: (1) the ZIKV NS2B-NS3 protease (NS2B-NS3pro) cleaves and inactivates Sox2, the key neural precursor transcription factor, and (2) ZIKV exploits multiple cellular genes to support its propagation in neural precursors. These events, especially when combined, cause neural stem/precursor cell apoptosis and reduce neurogenic capacity of surviving cells, ultimately leading to microcephaly in the fetus brain. Our interconnected and sharply focused Specific Aims are as follows: (1) Determine the physiological significance of NS2B-NS3pro-mediated proteolysis of Sox2 in the infected human neural stem/precursor cells, and (2) Identify cellular genes and major regulatory pathways that may lead to microcephaly in the infected newborns. To accomplish this aim, we will employ a plethora of the relevant cell-based assays, animal models and the advanced proteomics- transcriptomics-bioinformatics methodologies. The major outputs will be a body of knowledge that will be made publicly available and facilitate the next steps to disease eradication. Finally, it is worth reiterating that many flaviviruses cause serious neurological syndromes, and that our approach should be readily adaptable to other members of the extensive flaviviral family.